The purpose of this project is to determine the mechanisms responsible for interleukin-12's effectiveness as a mucosal adjuvant for protection against viral and bacterial infection. Our previous results have shown that intranasal treatment of mice with antigen and IL-12 enhances serum and respiratory antibody responses, and augments protection against influenza virus and Streptococcus pneumoniae. IgA appears to play an important role in IL-12-mediated protection and also in regulating inflammatory "danger" signals induced by IL-12. Macrophages from IgA-deficient mice over-express inflammatory cytokines upon stimulation. Furthermore, in vitro IgA treatment of murine macrophage cell lines inhibits production of TNF-a, IL-1, IL-6, nitric oxide, and nitric oxide synthase by these cells. The inhibitory effect appears to involve suppression of NF-KB activity after interaction of IgA with the macrophage cell surface. The experiments in this study will now investigate in detail the ability of IgA to control in vivo inflammation. The susceptibility of IgA-/- mice to inflammation will be examined after repeated inoculation of IL-12 +/- LPS. Induction of inflammation will be assessed by survival analysis, expression of inflammatory cytokines and histological examination of various tissues. The ability of exogenous IgA to prevent toxicity or ameliorate pre-existing inflammation will also be tested in mice by IgA inoculation during disease induction. The mechanism for protection will be analyzed in strains lacking the FcR common gamma chain, which is responsible in humans for mediating IgA cell signaling, and in strains lacking the polymeric IgR, which are unable to transport IgA across epithelial cell barriers into mucosal tissues. Finally, we will determine the intracellular signaling pathways that are responsible for inhibition of NF-KB activity and inflammatory cytokine expression after IgA binding to macrophages. Particular attention will be placed on the possibility that IgA recruits phosphatase enzymes in analogy to B, T, and NK cell inhibitory receptors. The results of this study will provide important insight into the roles of IL-12 and IgA in controlling immunity, and could have immediate impact on human health.